A robust immunoassay for pregnancy-associated plasma protein-A2 based on analysis of circulating antigen: establishment of normal ranges in pregnancy.

Abstract

Pregnancy-associated plasma protein-A (PAPP-A) and PAPP-A2, two homologous metzincin metalloproteases, are both tightly linked to regulation within the insulin-like growth factor (IGF) system because of their specific cleavage of IGF binding proteins. Recent studies suggest that PAPP-A may be involved in clinical conditions related to unwanted cellular growth, and the circulating levels of PAPP-A is an established biomarker in prenatal screening for chromosomal abnormalities. Microarray data indicate that PAPP-A2 has potential as a biomarker for pre-eclampsia. However, well-characterized immunological methods of quantification are not available. We therefore developed monoclonal antibodies against recombinant PAPP-A2. The antibodies were epitope mapped against recombinantly expressed chimeras between PAPP-A2 and PAPP-A. Furthermore, circulating PAPP-A2 was immunoaffinity purified and characterized by sequence analysis and mass spectrometry. Unlike PAPP-A, PAPP-A2 is a noncovalent dimer in which each subunit of 1558 amino acids originates from all of the 22 predicted coding exons. A previously hypothesized variant (PAPP-E) does not exist, but low amounts of a C-terminally truncated PAPP-A2 variant was detected. A sensitive and robust ELISA for full-length PAPP-A2 was developed and used to establish normal ranges of PAPP-A2 through pregnancy. The functional sensitivity of this ELISA at 20% CV was 0.08 ng/ml, and the serum concentration of PAPP-A2 was found to increase during pregnancy in agreement with placental synthesis. The existence of this assay will enable an assessment of the biomarker potential of PAPP-A2 in pre-eclampsia as well as other clinical conditions.

• Authors: S Kløverpris, E Gaidamauskas, L C V Rasmussen, M T Overgaard, C Kronborg, U B Knudsen, M Christiansen, A Kumar, C Oxvig
• Journal: Molecular human reproduction (Mol Hum Reprod) Vol. 19 Issue 11 Pg. 756-63 (Nov 2013) ISSN: 1460-2407 [Electronic] England
• PMID: 23804707 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)

Chemical References

• Antigens
• Biomarkers
• PAPPA2 protein, human
• Pregnancy-Associated Plasma Protein-A

Topics

• Animals
• Antigens (blood)
• Biomarkers (blood)
• Female
• Fetal Diseases (blood, diagnosis)
• HEK293 Cells
• Humans
• Immunoassay (methods)
• Mice
• Mice, Knockout
• Pre-Eclampsia (blood, diagnosis)
• Pregnancy (blood)
• Pregnancy-Associated Plasma Protein-A (analysis)
• Prenatal Diagnosis (methods, standards)
• Reference Values
• Sensitivity and Specificity